About a quarter of patients with severe aplastic anemia remain pancytopenic despite immunosuppressive therapy, and others have suboptimal responses in at least one lineage. We have developed a program to ask whether use of a drug stimulating the c-mpl receptor, which binds the endogenous hormone thrombopoietin, could stimulate human hematopoietic stem cells in vivo. Murine, our own rhesus, and human data all suggest that thrombopoietin can stimulate primitive hematopoietic stem and progenitor cells to expand. The small molecule oral thrombopoietin mimetic eltrombopag was initially developed to overstimulate platelet production from marrow megakaryocytes to compensate for immune platelet destruction. We developed a protocol utilizing eltrombopag in a phase 1/2 clinical trial for patients with refractory severe aplastic anemia. We initially reported that eltrombopag had efficacy in this setting with 44% (11/25) of patients having clinically significant hematologic responses (Olnes et al, NEJM 2012). We then reported additional safety and efficacy data on an expanded cohort of 43 patients, confirming an overall response rate or 40% with 3 to 4 months of treatment, including tri- and bilinear responses (Desmond et al, Blood, 2014). The majority of patients who remained on eltrombopag in an extension study (14/17) continued to show improvement, and 7 eventually had significant increases in neutrophil, red cell, and platelet lineages. Five patients with robust near-normalization of blood counts had drug discontinued at a median of 28.5 months after entry (range, 9-37 months), and all maintained stable counts a median of 13 months (range, 1-15 months) off eltrombopag. Eight patients, including 6 nonresponders and 2 responders, developed new cytogenetic abnormalities on eltrombopag, including 5 with chromosome 7 loss or partial deletion. This program resulted in FDA approval of a new labeled indication for eltrombopag in August 2014, and European commission approval in late 2015 for eltrombopag treatment of refractory aplastic anemia. This is the first new drug approved for aplastic anemia in decades and the first drug approved specifically for the refractory aplastic anemia patient population. During the current reporting period, we focused on an additional study, asking whether more prolonged administration of eltrombopag to patients with refractory SAA, for 6 months instead of 3 months, would improve response rate and rescue a larger fraction of refractory patients. This protocol is ongoing and has completed accrual. To date, responses appear to be faster than on the initial trial, due to starting at full dose, in contrast to dose escalation in the original trial. The overall response rate appears similar, although there were patients that responded between 3 and 6 months, who would have been deemed non responders in the first trial. We have combined both cohorts for molecular analyses asking whether EPAG impacts on clonal progression, and whether HPSC carrying mutations in specific genes linked to myeloid malignancies or other clones defined by new somatic mutations are specifically stimulated by EPAG treatment. Whole exome sequencing was performed in baseline samples and 6 month samples in all responding patients, and in any patient that developed cytogenetic abnormalities while on study. We found no evidence for specific clonal expansions related to EPAG treatment. In collaboration with Drs Neal Young and Danielle Townsley, we completed a trial adding EPAG to ATG and cyclosporine for the initial treatment of severe aplastic anemia. This trial was published (#1) and demonstrated a higher rate of complete responses at 6 months compared to historical controls, faster responses, and a higher overall rate of response in the patients that received EPAG. This trial is detailed in Dr Young's report. In addition, we are collaborating on a trial utilizing EPAG in patients with moderate aplastic anemia. This trial has completed accrual and analysis is in progress, showing a substantial response rate. In collaboration with Dr Andre Larochelle, we have elucidated the likely mechanism of EPAG activity in patients with high endogenous levels of TPO. This work will be described in his annual report